Robert et al. Molecular Neurodegeneration (2020) 15:23 https://doi.org/10.1186/s13024-020-00366-8 RESEARCH ARTICLE Open Access Cerebrovascular amyloid Angiopathy in bioengineered vessels is reduced by high- density lipoprotein particles enriched in Apolipoprotein E Jerome Robert1,2,3* , Emily B. Button1,2, Emma M. Martin1,2, Luke McAlary2,4, Zoe Gidden2, Megan Gilmour1,2, Guilaine Boyce1,2, Tara M. Caffrey1,2, Andrew Agbay1,2, Amanda Clark1,2, Judith M. Silverman2,5, Neil R. Cashman5 and Cheryl L. Wellington1,2,6,7 Abstract Background: Several lines of evidence suggest that high-density lipoprotein (HDL) reduces Alzheimer’s disease (AD) risk by decreasing vascular beta-amyloid (Aβ) deposition and inflammation, however, the mechanisms by which HDL improve cerebrovascular functions relevant to AD remain poorly understood. Methods: Here we use a human bioengineered model of cerebral amyloid angiopathy (CAA) to define several mechanisms by which HDL reduces Aβ deposition within the vasculature and attenuates endothelial inflammation as measured by monocyte binding. Results: We demonstrate that HDL reduces vascular Aβ accumulation independently of its principal binding protein, scavenger receptor (SR)-BI, in contrast to the SR-BI-dependent mechanism by which HDL prevents Aβ- induced vascular inflammation. We describe multiple novel mechanisms by which HDL acts to reduce CAA, namely: i) altering Aβ binding to collagen-I, ii) forming a complex with Aβ that maintains its solubility, iii) lowering collagen- I protein levels produced by smooth-muscle cells (SMC), and iv) attenuating Aβ uptake into SMC that associates with reduced low density lipoprotein related protein 1 (LRP1) levels. Furthermore, we show that HDL particles enriched in apolipoprotein (apo)E appear to be the major drivers of these effects, providing new insights into the peripheral role of apoE in AD, in particular, the fraction of HDL that contains apoE. Conclusion: The findings in this study identify new mechanisms by which circulating HDL, particularly HDL particles enriched in apoE, may provide vascular resilience to Aβ and shed new light on a potential role of peripherally-acting apoE in AD. Keywords: Alzheimer’s disease, Cerebrovasculature, High-density lipoprotein (HDL), Cerebral amyloid angiopathy (CAA), Endothelial inflammation, Apolipoprotein (apo)E, Tissue engineering * Correspondence: [email protected] 1Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada 2Djavad Mowafaghian Centre for Brain Health, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Robert et al. Molecular Neurodegeneration (2020) 15:23 Page 2 of 21 Background serum HDL-cholesterol (> 55 mg/dl) in cognitively normal As our population ages, the burden of cardiometabolic, elderly individuals associates with reduced AD risk (hazard neurodegenerative and neuroinflammatory diseases is ratio(HR)0.4)afteradjustingforAPOE genotype and growing rapidly. Alzheimer’s Disease (AD) is defined by vascular risk factors [21]. In mouse models of amyloidogen- the presence of beta-amyloid (Aβ) deposits and neuro- esis, deficiency of apoA-I, which leads to low HDL levels, is fibrillary tangles in the brain and affects over 50 million reported to selectively exacerbate CAA and cerebrovascular people worldwide with a global economic burden closed inflammation [22, 23], although other groups recently re- to one trillion US dollars [1]. Despite decades of re- ported either no effect [24]ortheoppositeresult[25]. search, there is not yet an approved disease-modifying Conversely, genetic apoA-I overexpression reduces CAA therapy for AD, as many drugs that target parenchymal and neuroinflammation [26], and delivery of recombinant pathologies have been ineffective in attenuating cognitive HDL or apoA-I Milano into the systemic circulation acutely decline [2]. One exception may be the anti-amyloid im- decreases soluble brain Aβ levels and leads to long- munotherapy aducanumab, which clears amyloid from lasting lowering of CAA and neuroinflammation, respect- the brain [3] and appears to slow cognitive decline at the ively [27, 28]. Importantly, although lipoprotein composition highest dose so far tested [4]. However, amyloid related is generally similar between mice and humans [29], circulat- imaging abnormities (ARIA), including edema (ARIA-E) ing lipids are mainly carried by HDL in rodents whereas and microhemorrhage (ARIA-H) are adverse effects they are mainly carried on low density lipoproteins (LDL) in associated with anti-amyloid immunotherapies and these humans, which may limit the translational relevance of some risks are elevated in elderly people with cerebrovascular animal model studies [30]. disease [5]. Thus, novel preventative and therapeutic We previously described a new human in vitro model of approaches are urgently needed, particularly with respect three-dimensional (3D) perfusable bioengineered vessels to the vascular contributions to AD. to study CAA and Aβ-associated vascular inflammation The importance of the cerebrovasculature in AD is [31, 32]. In this study we use this vascular model to define underscored by the observation that up to 90% of AD aut- several mechanisms by which HDL reduces Aβ accumula- opsy cases have evidence of cerebral small vessel disease, tion in the engineered vascular wall and attenuates Aβ- including vascular microbleeds, vascular degeneration and induced vascular inflammation. First, the anti-CAA and deposition of Aβ in the vascular wall, a condition known as anti-inflammatory functions of HDL are mediated by cerebral amyloid angiopathy (CAA) [6]. Sporadic CAA is distinct mechanisms. HDL prevents Aβ vascular accumu- also present in 10–40% of elderly brains [7]. Cerebrovascu- lation independently of its primary receptor, scavenger lar pathologies precede other pathophysiological conditions receptor (SR)-BI, in contrast to the previously described and are believed to exacerbate or even initiate AD and neu- SR-BI-dependent mechanism by which HDL prevents Aβ- roinflammation [8], perhaps by reflecting an age-related induced monocyte binding [32]. Second, delving deeper decline in the ability to clear Aβ from the brain by multiple into HDL’s anti-CAA functions, we define four distinct pathways involving the cerebrovasculature [9]. Addition- pathways that HDL uses to attenuate Aβ accumulation, ally, multiple cardiovascular risk factors such as hyperten- namely: i) altering Aβ binding to collagen-I, ii) forming a sion, type II diabetes and dyslipidemia increase AD risk complex with Aβ that maintains its solubility, iii) dimin- [10, 11] and a favourable cardiovascular health or manage- ishing collagen-I protein levels produced by smooth ment of cardiovascular risk factors, especially at midlife, muscle cells (SMC), and iv) attenuating Aβ uptake into may attenuate dementia risk decades later [12–14]. The SMC that is associated with reduced low density lipopro- clinical precedence of systemic factors that can affect AD tein receptor-related protein (LRP)1 levels. We also define pathogenesis raises the possibility that promoting vascular a specific subfraction of HDL that may be particularly resilience may be an attractive preventative or therapeutic relevant to AD. Although only ~ 6% of circulating HDL approach [15, 16]. lipoprotein particles contain apoE [33], it is this fraction of High density lipoprotein (HDL) is a circulating lipid apoE-containing HDL (HDL-apoE) that appears to have carrier that is best known for its pivotal role in reverse the most potent effects on vascular amyloid deposition. cholesterol transport, which is the process by which ex- ApoE is a major genetic risk factor for sporadic AD, cess cholesterol is removed from cells and transported where, in the brain, apoE is produced by astrocytes, to the liver for excretion into bile [17]. HDL also has microglia, pericytes and stressed neurons and contributes to potent anti-thrombotic, anti-oxidant, anti-inflammatory AD pathogenesis by modulating multiple pathways, includ- and cytoprotective functions [18], all of which may affect ing but not limited to Aβ metabolism, tauopathy, and neuro- AD. There is epidemiological evidence that AD risk can inflammation [34]. Our observations provide new evidence be attenuated by HDL levels [19]. Specifically, apolipo- that peripheral apoE, particularly the apoE found on a small protein (apo)A-I, the major HDL-associated protein, fraction of circulating HDL lipoprotein particles, may play a positively correlates with cognitive scores [20] and high greater role in AD pathogenesis than previously recognized. Robert et al. Molecular Neurodegeneration (2020) 15:23 Page 3 of 21 Methods Fabrication of engineered vessels Cells, lipoproteins and bioengineered tissues Bioengineered vessels were fabricated using a dynamic,